Role of genetic variants in cancer-associated fibroblasts in gastrointestinal cancers

• Applicant: Yoshiaki Sunami, Ph.D.
• Subject Area: Gastroenterology
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 534195368

The tumor microenvironment plays a key role in the progression and drug resistance of cancer. A variety of cellular as well as non-cellular components including immune cells, cancer-associated fibroblasts (CAFs), and extracellular matrix (ECM) compose the complex tumor microenvironment. CAFs are the major stromal cells and source of ECM proteins. Somatic single nucleotide variants (SNVs) activate downstream signaling pathways that are widely recognized as a hallmark of cancer. Yet, SNVs in CAFs and their respective functional consequence on proliferation and differentiation of CAFs, as well as tumor initiation, progression, and metastasis have been rarely demonstrated. To that end, we performed whole-exome sequencing of genomic DNA of isolated primary CAFs from pancreatic cancer patients. We have successfully identified SNVs in pancreatic CAFs, and shown that a SNV in gene called SERPINB2 can result in a truncated protein thereby leading to full or partial loss-of-function of this putative tumor suppressor which may enhance tumor progression. Among the other identified genes, four genes namely CNTNAP4, DENND4B, MIGA2 (FAM73B), and TOP2B exhibit association between expression and pancreatic cancer patient survival. Interestingly, consistently for all four genes, low expression of the gene is associated with shorter overall survival of pancreatic cancer patients. However, the role of these factors in the development and progression of gastrointestinal cancer has been barely elucidated. In this project, we therefore aim to 1) elucidate functional and pathological consequence of previously identified and four selected CAF-associated SNVs in pancreatic fibroblasts, and 2) investigate the impact of the respective SNV on CAF-immune-tumor cell interaction and its role on tumor development, progression, and invasion. Further, we aim to 3) identify stromal SNVs in several GI cancer types. To that end, we recruit patients for our project and collect GI cancer specimen. We will isolate CAFs from the resected material by outgrowth method and conduct exome sequencing and subsequent Sanger sequencing. To identify the role of pancreatic CAFs in signaling and molecular mechanisms, cancer development, progression and invasion, we will establish genome edited fibroblast cell lines using cutting-edge CRISPR tools. We will analyze protein expression, gene expression, signaling pathway activities, and paracrine factor production in these engineered fibroblast cell lines. Furthermore, we plan to establish organoid and co-culture models using pancreatic cancer cells and fibroblast cell lines, and we will examine the effects of stromal SNVs on pancreatic cancer phenotypes. With the data gained from the proposed project, we aim to establish a novel therapeutic strategy for gastrointestinal cancer patients by targeting stromal signaling pathways and paracrine factors to more effectively treat cancer patients in the future.

• DFG Programme: Research Grants
• Co-Investigator: Professor Dr. Jörg Kleeff